4B). Mice immunized with GFP+ CD8α+ cDCs from non-protected mice had equivalent bacterial titers as non-transferred animals upon challenge infection. In fact, only GFP+ CD8α+ cDCs from mice immunized with the protective dose of secA2−Lm were

able to induce substantial levels of immunity. Since the number of bacteria per infected cell is the same between the two conditions of immunization, it suggested that other signals distinct from those given by cytosolic bacteria are allowing CD8α+ cDCs from protected animals to be optimally conditioned to induce CD8+ T-cell protective memory. Protected mice were immunized with ten-fold more bacteria than non-protected click here animals, likely leading to a stronger inflammatory environment at the time of DC maturation. To provide support for this hypothesis, we measured the early inflammatory environment (5 h) under Seliciclib price the two conditions of immunization (Fig. 5). As proposed, we readily detected a strong inflammatory response

that included cytokines and chemokines involved in DC maturation in mice that received 107secA2−Lm. Animals injected with the lower numbers of bacteria were comparable to non-immunized control groups and exhibited low levels of inflammation. We next sought to determine whether this finding held true for animals immunized with other well-established Tangeritin protective Lm immunizations, e.g. wt Lm or the attenuated mutant actA−Lm25 (Supporting Information Fig. 5) and monitored several inflammatory mediators (IL-1β, CCL2, IL-12p70 and TNF-α) over a 48 h kinetics. In all groups that received protective immunization (e.g. 107secA2 Lm−, 106actA−Lm

and 3000 wt Lm), inflammation reached levels that were never measured in mice immunized with the non-protective dose of secA2−Lm. In the case of wt Lm immunization, however, such levels of inflammation were only observed at later time points (24–48 h), a result in agreement with former studies 26, which also correlates with the low initial inocula and the growth kinetics of wt Lm in vivo 16. Therefore, collectively these data favor the idea that during a protective immunization, CD8α+ cDCs receive stronger extracellular inflammatory signals than during non-protective immunization, which likely contribute to their optimal maturation in vivo. To further support to our interpretation that both cytosolically delivered and extracellular signals are conditioning CD8α+ cDC optimal programming, we compared the maturation profiles of infected and non-infected CD8α+ cDCs from mice immunized with the two doses of secA2−Lm.